Interleukin-21 (IL-21) is a type I cytokine that has been shown to modulate functions of T, B, NK, and myeloid cells, particularly in murine models. The IL-21 receptor (IL-21R) is closely related to the IL-2 receptor and is capable of transducing signals through its dimerization with the common gamma chain (3c). Our initial studies in peripheral blood lymphocytes of HIV+ subjects revealed two intriguing aspects of IL-21R signaling. First, it selectively upregulates expression of the cytolytic molecule perforin in CD8 T cells in vitro without inducing proliferation, unlike other cytokines that signal through 3c receptors. Second, the cells of aviremic HAART treated HIV infected persons were more responsive to IL-21 than cells of healthy HIV-uninfected volunteers. The present proposal will examine the molecular mechanisms involved in perforin upregulation by IL-21 in HIV+ versus HIV negative individuals and in its unique absence of inducing proliferative signals. Furthermore it will test the hypothesis that the perforin enhancing capabilities of IL-21 will augment cytolytic activity of CD8 T cells and Natural Killer (NK) cells against SIV in rhesus macaques. Our first aim is to extend previous observations and investigate the activity of IL-21 on T cells and NK cells of viremic and aviremic HIV infected patients for expression of perforin and IL-21R on T cells, NK cells, synthesis of IL- 21 by CD4 T cells and plasma IL-21 levels. In our 2nd aim in vivo studies in rhesus macaques are proposed to identify the dose at which IL-21 is safe and biologically active, to verify that it potentiates antiviral immune responses and to establish its effect on viral replication. For this aim we will be collaborating with Drs. Francois Villinger at Emory University for studies in healthy uninfected monkeys and Genoveffa Franchini at the National Cancer Institute for studies in monkeys chronically infected with the Simian immune deficiency virus (SIV). Our 3rd aim is to examine in detail the molecular mechanism of IL-21/IL-21R signaling resulting in perforin upregulation without causing proliferation. The role of Stat5 versus Stat3 and role of HIV-associated immune activation in promoting IL-21 signaling will be examined. These studies will guide future development of IL-21 for immunotherapy in HIV infection. Much progress has been made in the treatment of HIV-infected patients with potent antiretroviral drugs, but long term therapy carries the risk of drug related complications. There are also some patients who fail to control the virus despite treatment with potent drugs. This project is investigating interleukin-21 (IL-21), a biologic protein (cytokine) made naturally by the host which was found to increase perforin, a molecule used by killer cells to kill virally infected cells. The proposed studies will investigate the molecular basis of the action of IL-21 and test it for activity by infusing it in monkeys. If the results are promising, IL-21 could potentially be developed as an agent that could either augment immune responses against the AIDS virus or be used with vaccines to boost vaccine activity.